ATP binding cassette transporter A1 (ABCA1) is a membrane protein that is required to remove excess lipid from macrophages. Recent data have suggested that macrophage ABCA1 expression is anti-inflammatory and protects against development of insulin resistance. However, since Abca1 is expressed in nearly all tissues in the body, it is difficult to determine the specific role of macrophage Abca1 with regard to inflammation, atherosclerosis development, and insulin resistance. To address these gaps in knowledge, we have developed a macrophage-specific Abca1 knockout (MSKO) mouse. Our goal is to use these mice to determine the mechanisms by which macrophage-specific deletion of Abca1 expression: 1) increases macrophage inflammation, 2) affects atherosclerosis development, and 3) increases development of obesity and insulin resistance. We have found that macrophages from MSKO mice are hypersensitive to Toll-like receptor (TLR) agonists, which appears related to an increase in membrane free cholesterol and lipid rafts. In Specific aim 1, we hypothesize that macrophages from MSKO mice have an i , resulting in increased TLR and adapter protein recruitment into lipid rafts, increased signaling efficiency, , and a pro-inflammatory state. 2) the distribution of TLR4 and adapter proteins in lipid raft vs. non-raft membrane regions 1 TLR4 stimulation. Based on preliminary studies, we hypothesize in Specific aim 2 that atherosclerotic lesions in MSKO mice in the low density lipoprotein receptor (LDLr) KO background have fewer macrophages that are CE-enriched relative to LDLrKO mice, resulting in a similar degree of atherosclerosis. We will determine: 1) plasma lipoprotein phenotype, 2) in vivo reverse cholesterol transport from macrophages, 3) atherosclerosis extent, 4) aortic lesion macrophage number, lipid content, and gene expression, and 5) effect on MSKO-LDLrKO and LDLrKO mice fed an atherogenic diet. In Specific aim 3, our preliminary data demonstrated that MSKO mice fed a high fat/high sucrose (HF/HS) diet have increased body weight, increased hepatic lipid content, and systemic insulin resistance compared to WT mice. We hypothesize that the HF/HS diet results in pro-inflammatory macrophages that reduce insulin signaling and increase fat storage in peripheral tissues and liver. Using HF/HS diet-fed WT and MSKO mice ', we will determine plasma lipid/lipoprotein phenotype, macrophage inflammatory state and infiltration into tissues, whole body metabolic phenotype, and insulin signaling in liver and peripheral tissues. Results from these studies will fill gaps in knowledge on the specific role of macrophage-specific Abca1 expression in vivo in the pathogenesis of chronic inflammatory diseases such as atherosclerosis, insulin resistance and obesity. PUBLIC HEALTH RELEVANCE: Chronic inflammation results in obesity and insulin resistance. In this proposal, we seek to understand the relationship between macrophage inflammation that results from the specific deletion of a membrane cholesterol transporter on development of atherosclerosis, obesity and insulin resistance.